Reference to any prior art in this specification is not, and should not be taken as, an acknowledgement or any form of suggestion that this prior art forms part of the common general knowledge in Malaysia or any other country.
Bibliographic details of the publications referred to by author in this specification are collected at the end of the description.
The demand for oils and fats is expected to increase dramatically with the increase in world population and the need for sustainable resources. Oil palm plants, Elaeis guineensis and Elaeis oleifera, produce palm oil and palm kernel oil and represent the highest yielding oil crop in the world. Palm oil has been forecasted to contribute to around a quarter of the world's oil and fats demand by the year 2020 (Rajanaidu and Jalani, World-wide performance of DXP planting materials and future prospects. In Proceedings of 1995 PORIM National Oil Palm Conference. —Technologies in Plantation, The Way Forward. 11-12 Julai, Kuala Lumpur: Palm Oil Research Institute of Malaysia:1-29, 1995). Due to the demand, there is a need to increase the quality and yield of palm oil and palm kernel oil and to rapidly develop new characteristics when required.
Oil palm is the most important commercial crop in Malaysia. It has been identified as the most likely candidate for the development of a large scale production and renewal plant (Ravigadevi et al, Genetic engineering in oil palm. In Advances in Oil Palm Research. (eds). Yusof, Jalani, and Chan Malaysia Palm Oil Board. 1:284-331, 2000) for palm oil-derived chemicals. The ultimate aim is to genetically engineer the oil palm so as to modify its oil composition in order to expand its applicability. Moreover, the advancement in the genetic transformation of plants has made it possible to transfer foreign genes into the genome of oil palm (Parveez, PhD. Thesis. Universiti Putra Malaysia, 1998). Introduction of foreign genes via genetic engineering would potentially enhance the productivity and value of oil palm.
Genetic engineering can be used to improve plant quality by introducing and expressing selected genes into the plant genome. Genetic engineering processes are often unique to particular plants. An efficient tissue culture system is required in order to produce genetically modified plants after successful delivery and integration of genetic material into cells which regenerate into a whole plant. The introduced genetic material needs to be present in all cells of the regenerated plant and its progeny. The expression of the genetic material is controlled by promoters which may be constitutive or inducible, and operable ubiquitously in all plant cells or operable in a tissue specific manner.
A promoter is a short DNA sequence, usually upstream (5′) to the relevant coding sequence, to which RNA polymerase binds before initiating transcription. This binding aligns the RNA polymerase so that transcription will initiate at a specific site. The nucleotide sequence of the promoter determines the nature of the enzyme that attaches to it and the rate of RNA synthesis. The RNA is processed to produce messenger RNA (mRNA) which serves as a template for translation of the RNA sequence into the amino acid sequence of the encoded polypeptide. The 5′ non-translated leader sequence is a region of the mRNA upstream of the coding region that may play a role in initiation and translation of the mRNA. The 3′ transcription termination/polyadenylation signal is a non-translated region downstream of the coding region that functions in the plant cells to cause termination of the RNA and the addition of polyadenylate nucleotides to the 3′ end. It has been shown that certain promoters are able to direct RNA synthesis at a higher rate than others.
In order to functionally express a gene in plant, the transgene must have a promoter that is recognized by RNA polymerase in plant cells. There are two major classes of promoter, constitutive and regulated (or inducible). Constitutive promoters direct expression in virtually all tissues and are largely, if not entirely, independent of environmental and developmental factors. As their expression is normally not conditioned by endogenous factors, constitutive promoters are usually active across species and even across kingdoms. Tissue-specific or development-stage-specific promoters (regulated) direct the expression of a gene in specific tissue(s) or at certain stages of development. For plants, promoter elements that are expressed or affect the expression of genes in the vascular system, photosynthetic tissues, tubers, roots and other vegetative organs, or seeds and other reproductive organs can be found in heterologous systems (e.g. distantly related species or even other kingdoms) but the most specificity is generally achieved with homologous promoters (i.e. from the same species, genus or family). This is probably because the coordinate expression of transcription factors is necessary for regulation of the promoter's activity.
Because promoters affect transcription both quantitatively and qualitatively, the success of gene transfer technologies, varying from basic research to crop improvement and biopharming, depends on their efficacious selection and use (Potenza et al, In Vitro Cell. Dev. Biol. Plant 40:1-22, 2004). Plant promoters that are capable of driving high and constitutive expression of a particular transgene have become a valuable tool in plant genetic engineering. These promoters are required to select transgenic cells or plants that have a high level of production of the specific protein of interest. The high expression of a selectable marker is important to identify a non-transformed cell and to enable selection of a resistant transformant which will survive and generate into transgenic plant (Parveez, 1998, supra). This prevents non-transformant domination of the culture and promotes the growth of chimeric plants (Christou, The Plant J. 2:275-281, 1992; Ritala et al, Plant Mol. Biol. 24:317-325, 1994). The high level expression of a reporter protein such as GUS, GFP and CAT in a plant cell can also be achieved by using constitutive promoters. Moreover, the use of constitutive promoters is essential in the production of compounds that require ubiquitous activity in the plant and during all stages of plant development.
There are diverse spectra of constitutive promoters which are available for use in the genetic engineering of plants (Xiao et al, Molecular Breeding, 15:221-231, 2005). A commonly used example which results in the strong constitutive expression of a transgene in a plant is the CaMV35S promoter which originates from the cauliflower mosaic virus (Potenza et al, 2004 supra). Additionally, it has been reported that a number of widely used constitutive promoters include maize ubiquitin (Christensen and Quail, Transgenic Research 5:213-218, 1996), rice Actin 1 (McElroy et al, Plant Cell. 2:163-171, 1990) and maize derived Emu (Last et al, Theor. Appl. Genet., 81:581-588, 1991). Although a number of constitutive promoters have been isolated, it is still important to identify new plant promoters, especially for the high level expression of transgenes or other genetic material in selected plants or selected parts of plants. A wider range of effective promoters would make it possible to introduce multiple transgenes into plant cells while avoiding the risk of homology-dependent gene silencing (Xiao et al, 2005 supra).
As in other genetically engineered plants, constitutive promoters have previously been used to drive the expression of genes in transgenic oil palms. The promoters, Emu and Ubi1, were found to be the most efficient promoters in driving high level expression of foreign genes in oil palm plants (Chowdhury et al, Plant Cell Rep.:16:277-82, 1997). However, these promoters originated from unrelated plant species. Previous studies have shown that promoters can be more effective if isolated from the same species as the transgenic plant. McElroy et al, 1990 supra found, for example that β-glucuronidase (GUS) expression under the control of a rice actin promoter (Act1) in transformed rice protoplasts was approximately 6-fold greater than expression induced under the control of the maize Adh1 promoter.
There is a need to identify promoters derived from the oil palm genome to facilitate genetic manipulation of oil palm plants as well as other plant species.